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. 2021 Nov 26;24(12):103517. doi: 10.1016/j.isci.2021.103517

Figure 3.

Figure 3

GFAT2-Akt axis positively regulates ISO-induced cardiomyocyte hypertrophy

(A) Representative immunoblots of Akt, phospho-Akt(S473), ERK, phospho-ERK, p38, phospho-p38, JNK, phospho-JNK, and GAPDH in NRVMs treated with indicated small interfering RNA (siRNA) in the presence or absence of ISO (1 μM, 12 h).

(B–E) Quantitative analysis of phospho-Akt(S473), phospho-ERK, phospho-p38, phospho-JNK in NRVMs treated with indicated siRNA in the presence or absence of ISO (1 μM, 12 h) (n = 6).

(F) Immunoprecipitation assays using lysates of NRVMs treated with indicated siRNA in the presence or absence of ISO (1 μM, 12 h) (n = 6). After immunoprecipitation with control IgG or an O-GlcNAcylation antibody, immunoblotting for Akt was performed. Immunoblots of input controls (10% lysates) are also shown.

(G) Representative immunoblots of Akt, phospho-Akt(S473), and GAPDH in NRVMs treated with adenovirus harboring wild-type Akt or mutant Akt in T479A in the presence of ISO (1 μM, 24 h). Quantitative analysis of phospho-Akt(S473) in NRVMs treated with adenovirus harboring wild-type Akt or mutant Akt in T479A in the presence of ISO (1 μM, 24 h).

(H) Cell surface area of NRVMs treated with adenovirus harboring wild-type Akt or mutant Akt in T479A in the presence of ISO (1 μM, 24 h) (n = 150 cells in each group). Scale bar, 50 μm.

(I) Cell surface area of NRVMs treated with GFAT2 siRNA and/or MK-2206(1 nM, 24 h) in the presence of ISO (1 μM, 24 h) (n = 150 cells in each group). Scale bar, 50 μm. IP, immunoprecipitation. ∗p < 0.05, ∗∗p < 0.01: post hoc Tukey's comparison test.